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Choosing Your Perfect Plasmid Backbone

Background

When choosing what plasmid backbone to use, you have many elements
to consider. Here is a guide to Addgene's empty vector backbones. For
the most part, we will assume that you want to express a gene;
however, we have a section at the end for if you are studying a
different genetic element or want to express shRNA.

Epitope Tag or Fusion Protein

Tags and fusion proteins are excellent tools for further
understanding the function of your favorite gene. For example, fusing
your protein to an epitope tag, such as Flag or HA, will allow you to
easily identify your protein using an antibody against that epitope.
This could allow you to conduct western blots or immunoprecipitations
of your favorite gene even if you do not have an antibody against it.
Another common scenario is fusing your protein to another protein, such
as GFP, which allows you to visualize the cellular localization of your
protein.

Just remember that when you are designing your plasmid you should
keep your gene "in frame" with the fusion protein. This means that the
final product should be translated as a single string of amino acids
that preserves the sequence of your gene and of the fusion protein.

Selectable Markers

Regardless of your delivery method, it's unlikely that all of your
cells will take up your plasmid. Thus, many plasmids have markers on
them so that you can find or select for only the cells that received
the plasmid.

Viral Expression and Packaging

Although transient expression is sufficient for some experiments,
scientists often want to create stable cell lines in which the plasmid
is incorporated into the host DNA. For mammalian cells, you can do this
through viral delivery. Visit our viral vector page for more information. Below are some common delivery methods:

Delivery
method

Advantages

Representative Empty
Backbones

Lentiviral

Can transduce both dividing and nondividing
cells

Addgene has an extensive collection of plasmids for
packaging and expression. See our dedicated lentiviral plasmid page.